Apoptosis is a cell suicide process of sequential biochemical events triggered by a variety of physiological and stress stimuli. Several lines of evidence indicate that a family of cysteine proteases, or caspases (Alnemri et al., 1996) play a crucial role in execution of apoptosis. Several members of the caspase family have been identified (for review, see Henkart, 1996; for nomenclature, see Alnemri et al., 1996) which share certain characteristic features. For example, all the identified caspases contain a conserved motif QAC(R/Q)G, in which the Cys residue is the structural hallmark of a caspase. This cysteine residue, together with two highly conserved residues, corresponding to His237 and Gly238 in ICE, may form the active site of a caspase (Wilson et al., 1994; Walker et al., 1994). In addition, many members of the family are capable of inducing apoptosis when overexpressed in mammalian cells (Henkart, 1996).
Many divergent stimuli can activate the caspase cascades leading to apoptosis. In recent years, apoptosis induced by TNF and FasL have received extensive attention. TNF elicits a broad range of biological effects (Goeddel, 1986; Beutler and Cerami, 1988; Fiers, 1991) through two distinct membrane receptors, TNF-R1 and TNF-R2, which are expressed at low levels on most cell types (Loetscher et al., 1990; Schall et al., 1990; Smith et al., 1990; Fiers, 1991; Tartaglia and Goeddel, 1992). Apoptosis induced by TNF is mediated primarily through TNF-R1. The intracellular domain of TNF-R1 contains a “death domain” of approximately 80 amino acids that is responsible for signaling cell death by the receptor (Tartaglia et al., 1993). A homologous death domain is also found in the cytoplasmic region of Fas and Ws1/DR3/Apo-3, two other members of the TNF receptor family that can potently induce apoptosis (Itoh and Nagata, 1993; Chinnaiyan et al., 1996b; Kitson et al., 1996; Marsters et al., 1996).
TRADD, a cytoplasmic protein containing a C-terminal death domain, interacts with the death domain of TNF-R1 in a ligand dependent process (Hsu et al., 1995; Hsu et al., 1996a). As observed for TNF-R1, overexpression of TRADD causes both apoptosis and activation of NF-κB (Hsu et al., 1995). The death domain of TRADD also interacts with the cytoplasmic protein FADD (Hsu et al., 1996a) through their respective death domains. Fas and FADD have also been shown to interact directly through their respective death domains (Boldin et al., 1995; Chinnaiyan et al., 1995). Although the death domains of TNF-R1, Fas, and TRADD induce apoptosis following overexpression in mammalian cells (Tartaglia et al., 1993; Hsu et al., 1995; Itoh and Nagata, 1993; Hsu et al., 1996b), overexpression of the C-terminal death domain of FADD inhibits TNF- and Fas-induced cell death (Chinnaiyan et al., 1996a; Hsu et al., 1996a). The N-terminal domain of FADD, termed death effector domain (DED), induces apoptosis after overexpression, suggesting the DED of FADD may activate a downstream cell death signaling component (Chinnaiyan et al., 1996a; Hsu et al., 1996a).
A recently identified a cysteine protease, caspase-8, (previously called Mch5, MACH and FLICE, Fernandes-Alnemri et al., 1996; Boldin et al., 1996; Muzio et al., 1996) may represent the missing link between FADD and the basic cell death machinery. The N-terminal domain of caspase-8 contains two DED-like modules through which it interacts with FADD (Boldin et al., 1996; Muzio et al., 1996). The C-terminal domain of caspase-8 is homologous to members of the caspase family and has protease activity towards most known caspases and PARP (Fernandes-Alnemri et al., 1996; Srinivasula et al., 1996; Muzio et al., 1996). The present invention provides a novel family of proteins termed Casper (caspase-eight-related protein), which are structurally related to caspase-8.